Chromosomal nonhistone proteins are degraded by chromosomal protease in urea and in mixtures of NaCl or guanidine-HCl and urea (at wide pH ranges) under conditions which are frequently used for fractionation of chromosomal nonhistone proteins and reconstitution of chromatin. The degradation of nonhistone proteins is not blocked by certain inhibitors of trypsin and chymotrypsin or by NaHSO3. The presence of proteases in chromatin preparations is a hinderance to evaluation of chromosomal proteins, and consequently these protease require detailed investigation. In this study we shall investigate the possible degradation of nonhistone proteins during storage of chromatin and during assay of template activity of chromatin. We shall study the possible presence of different chromosomal proteases for histones and nonhistone proteins. Also, we shall test various reported protease inhibitors for inhibition of the degradation of nonhistone proteins by chromosomal protease. If this approach does not produce effective inhibitors of the degradation of nonhistone proteins, we shall attempt the fractionation and characterization of proteases in the hope that information can be obtained for the rational design of effective inhibitors.